This invention is in the area of organic synthesis of nucleosides, and in particular relates to a process for the preparation of enantiomerically pure .beta.-D-(-)-dioxolane nucleosides.
A number of 2',3'-dideoxynucleosides have been found to be potent antiviral agents against human immunodeficiency virus (HIV), the causative agent of acquired immunodeficiency syndrome (AIDS). The lead compound, AZT (Mitsuya, H.; Broder, S. Proc. Natl. Acad. Sci. U.S.A., 1986 83, 1911) has been approved by the U.S. Food and Drug Administration for patients with AIDS and AIDS-related complex. Several other 2',3'-dideoxynucleosides are undergoing various stages of clinical trials, including 3'-azido-2',3'-dideoxyuridine (AZDU or CS-87, see, Chu, C. K.; et al., J. Med. Chem., 1989, 32, 612; and Eriksson, B. F. H.; et al., Antimicrob. Agents Chemother., 1989, 33, 1927), 2',3'-dideoxyinosine (DDI) and 2',3'-dideoxycytidine (DDC) (see Yarchoan, R. et. al., Science, 1989, 245, 412), 3'-deoxy-2',3'-didehydrothymidine (D4T, Lin, T. S., et al., Biochem. Pharmacol., 1987, 36, 311; Hamamoto, Y., et al., Antimicrob. Agents Chemother., 1987, 31, 907; Balzarini, J., et al., Biochem. Biophys. Res. Commun., 1987, 140, 735 ), and 2'-fluoro-arabinofuranosyl- 2'-3'-dideoxycytidine (Martin, T. A., et al., J. Med. Chem., 1990, 33, 2137; Watanabe, K. A., et al., J. Med. Chem., 1990, 33, 2145; Sterzycki, R. Z., et al., J. Med. Chem., 1990 33, 2150).
In the 5'-triphosphorylated form, these nucleosides are known to inhibit HIV reverse transcriptase as well as cause chain-termination of the growing viral DNA chain. Furman, P. A., et al., Proc. Natl. Acad. Sci. U.S.A., 1986, 83, 8333; Cheng, Y. C., e al., J. Biol. Chem., 1987, 262, 2187; St. Clair, M. H., et al., Antimicrob. Agents Chemother., 1987, 31, 1972; and Schinazi, R. F., et al., Antimicrob. Agents Chemother., 1989 33, 115.
The stereochemistry of nucleoside derivatives play an important role in their biological activity. The C1' position of the ribose in the nucleoside (the carbon bound to the nitrogen of the heterocyclic base) is a chiral center because the carbon is attached to four different moieties. Likewise, there is an optically active center at C4' of the nucleoside (the ring carbon bound to the hydroxymethyl group that is phosphorylated in nucleotides). In the naturally occurring nucleosides, both the base attached to the C1' and the hydroxymethyl group attached to the C4' atom are in the .beta.-configuration (above the plane of the sugar). The corresponding non-naturally occurring .alpha.-isomers (in which the moieties are below the plane of the sugar) are rarely biologically active, and are typically toxic.
An analysis of the solid-state conformations of six active and two inactive anti-HIV nucleoside agents was recently performed to attempt to correlate the presence or absence of certain stereochemical features with high HIV activity. Van Roey, P., et al., J. Am. Chem. Soc., 1988, 110, 2277; and Van Roey, P., et al., Proc. Natl. Acad. Sci. U.S.A., 1989, 86, 3929. The x-ray structures indicated that active anti-HIV nucleosides assume the C3'-exo or similar carbohydrate conformations while inactive compounds prefer the C3'-endo conformation. (Endo and exo refer to the conformations in which the atoms are at the same or opposite side of the sugar ring in relation to the base). The C3'-exo and C3'-endo conformations place the C5' atom in axial and equitorial positions, respectively. The position of the C5' atom affects the location of the 5'-hydroxyl group in relation to the base. Since the 5'-hydroxyl group is the site of phosphorylation of the nucleoside, its location with respect to the rest of the nucleoside is important.
There has been recent interest in the synthesis of nucleoside derivatives in which the 3'-carbon of the nucleoside has been replaced with a heteroatom. Norbeck, D. W., et al., in Tet. Lett., 1989, 30, 6263, reported the synthesis of (.+-.)-1-[(2.beta.,4.beta.)-2-(hydroxymethyl)-4-dioxolanyl]thymine (referred to below as (.+-.)-dioxolane-T, see FIG. 1), that results in a racemic mixture of diastereomers about the C4' atom. The product is a derivative of 3'-deoxythymidine in which the C3' atom has been replaced with an O3' atom. The product was synthesized in five steps from benzyloxyaldehyde dimethylacetal and (.+-.)-methyl glycerate to produce a 79% yield of the 1:1 diastereomeric mixture. The X-ray crystallographic analysis of the product revealed that the dioxolane ring adopts the .sup.3 T.sub.4 conformation commonly observed in ribonucleosides, with the O3' atom in the endo position. Norbeck reported that the racemic mixture of dioxolane-T exhibits an anti-HIV activity of 20 .mu.M in ATH8 cells, and attributed the low efficacy against the virus to an effect of the endo conformation of the O3' atom.
Belleau, et al., in the Fifth International Conf. on AIDS, Montreal, Canada June 4-9, 1990, paper No. T.C.O.1, reported a method of synthesis of cytidine nucleosides that contain oxygen or sulfur in the 3' position. The dioxolane ring was prepared by the condensation of RCO.sub.2 CH.sub.2 CHO with glycerine. As with the Norbeck synthesis, the Belleau synthesis results in a racemic mixture of diastereoisomers about the C4' carbon of the nucleoside. Belleau reported that the sulfur analog, referred to as NGBP-2I or (.+-.) BCH-189 (see FIG. 1), had high anti-HIV activity. (.+-.) BCH-189 is currently undergoing preclinical toxicology.
To date, no one has reported a method of synthesis of a nucleoside analog with an oxygen in the 3'-position that results in an enantiomerically pure dioxolane nucleoside that has the same stereochemistry as the nucleosides found in nature (the .beta. stereoisomer). There is a need for such a synthesis as a research tool to provide more information on the effect of stereochemistry on the anti-viral activity of nucleoside derivatives, and to provide new anti-HIV agents.
It is therefore an object of the present invention to provide a method of synthesis of enantiomericaly pure dioxolane nucleosides.
It is another object of the present invention to provide enantiomerically pure dioxolane nucleosides with significant anti-HIV activity.